For patients with non-small cell lung cancer (NSCLC), treatment response and disease progression are typically assessed based on radiographic changes in tumor size or metabolic activity. However, imaging studies can sometimes yield equivocal results that lead to confusion in clinical decision-making. For example, after radiation therapy for lung cancer, in-field tumor recurrence can be difficult to distinguish from radiation-induced pneumonitis or fibrosis based on radiographic appearance alone. Imaging can also miss small metastatic deposits after completion of curative-intent stereotactic body radiotherapy (SBRT) or concurrent radiation and chemotherapy for localized NSCLC. For patients treated with new immunotherapies, confusion commonly arises when trying to assess therapeutic efficacy because radiographic shrinkage of disease can be delayed by several months. For many other types of cancer, information from quantitative changes in circulating tumor markers can be used to supplement radiographic findings. Unfortunately, no such blood markers have been found to be routinely useful for lung cancer. In this proposal, we aim to investigate whether a novel, highly cancer-specific class of biomarkers - circulating tumor DNA (ctDNA) - might be well suited for this purpose. In particular, we will focus on clinical scenarios in that are commonly prone to radiographic ambiguity. Our laboratory is especially well equipped to carry out this study because we have developed a robust, ultrasensitive next-generation sequencing-based assay that allows broad-spectrum measurement of mutant ctDNA. We propose to take advantage of this novel technology to investigate the following Specific Aims: Aim 1: Establish estimates of ctDNA assay sensitivity and specificity in patients with early-stage, locally advanced, and metastatic NSCLC prior to initiating therapy. Aim 2: For patients treated with stereotactic body radiotherapy for inoperable early-stage NSCLC or with concurrent radiation and chemotherapy for locally advanced NSCLC, determine whether detectable or rising levels of ctDNA after completion of treatment correlate with disease recurrence. Aim 3: For patients treated with immunotherapy for metastatic NSCLC, determine whether an early drop in ctDNA levels can predict subsequent radiographic response.